Most fast excitatory synaptic transmission in the central nervous system is mediated by the neurotransmitter glutamate. Aberrant glutamatergic neurotransmission, most notably of the NMDA receptor subtype, has been implicated in a wide variety of nervous system diseases including psychiatric disorders (e.g. schizophrenia and mood disorders), chronic neurodegenerative diseases (e.g. multiple sclerosis and Parkinson's and Alzheimer's diseases), and disorders arising from acute excitotoxicity (e.g. epilepsy and stroke). Accordingly, there is a great need for pharmacological agents that target dysfunctional glutamatergic signaling without resulting in the alteration of its normal function or the occurrence of adverse side effects (e.g. drowsiness, hallucinations, and coma). There are presently few glutamatergic-based therapies available for clinical use, but those agents that are clinically useful (amantidine and memantine) share a common theme: they modulate NMDA receptor gating (opening/closing of the ion channel) in an activity-dependent manner. Thus, understanding gating mechanisms are critical to developing novel therapeutic agents. Glutamate receptors (GluRs) are tetramers being composed of identical or similar subunits. Recently, a crystal structure of a nearly intact AMPA receptor, composed of identical subunits, showed that the receptor subunits exist in two unique conformations (termed 'A/C' and 'B/D') with subunits of the same conformation being positioned opposite one another. Functional NMDA receptors are obligate heterotetramers predominantly comprised of two GluN1 and two GluN2 subunits that adopt the 'A/C' and 'B/D' conformations respectively. In addition, native GluR subtypes (AMPA and kainate) are thought to be heterotetramers composed of at least two different subtypes. The major goal of my proposal is to study the significance of these two conformations in heteromeric AMPA and kainate receptors (Aim 1) and its significance to glutamate receptor gating (Aim 2). To address these aims, I will take advantage of specific cysteine-substituted mutations in the M3-S2 linkers, a short polypeptide that connects the ligand-binding domain to the ion channel pore, to determine a subunit's general conformation through the use of immunoblots and functional assays (patch-clamp electrophysiology). To delineate the significance of distinct conformational states in receptor function, I will alter the physical length and electrostatic composition of the M3-S2 linkers in obligate heteromultimeric NMDA receptors. Through the use of varying agonists and antagonists, the resulting data will provide insights into the gating mechanisms underlying glutamatergic signaling and allow for the innovation and generation of specific pharmacological agents aimed to treat glutamate-based diseases. PUBLIC HEALTH RELEVANCE: Glutamate receptor dysfunction has been implicated in many pathological nervous system disorders including psychiatric, neurodegenerative, and neurodevelopmental diseases. Clinically few glutamate-based therapies exist due to their non-selective targeting of glutamate receptors and of these, the most promising agents target specific aspects of glutamate receptor gating. Thus, by defining novel aspects of glutamate receptor structure-function my studies will serve to further understand glutamate receptor gating and may lead to novel strategies to treat glutamate-based neurological diseases.